Interplay among NH4+ uptake, rhizosphere pH and plasma membrane H+-ATPase determine the release of BNIs in sorghum roots - possible mechanisms and underlying hypothesis

Aims and background The ability to suppress soil nitrification through the release of nitrification inhibitors from plant roots is termed 'biological nitrification inhibition' (BNI). Earlier, we reported that sorghum roots release higher BNI-activity when grown with NH4+, but not with NO3- as N source. Also for BNI release, rhizosphere pH of <= 5.0 is needed; beyond this, a negative effect on BNI release was observed with nearly 80% loss of BNI activity at pH >= 7.0. This study is aimed at understanding the inter-functional relationships associated with NH4+ uptake, rhizosphere-pH and plasma membrane H+-ATPase (PM H+-ATPase) activity in regulating the release of BNIs (biological nitrification inhibitors) from sorghum roots. Methods Sorghum was grown hydroponically and root exudates were collected from intact plants using a pH-stat system to separate the secondary acidification effects by NH4+ uptake on BNIs release. A recombinant luminescent Nitrosomonas europaea bioassay was used to determine BNI-activity. Root plasma membrane was isolated using a two-phase partitioning system. Hydrolytic H+-ATPase activity was determined. Split-root system setup was deployed to understand the localized responses to NH4+, H+-ATPase-stimulator (fusicoccin) or H+-ATPase-inhibitor (vanadates) on BNI release by sorghum. Results Presence of NH4+ in the rhizosphere stimulated the expression of H+-ATPase activity and enhanced the release of BNIs from sorghum roots. Fusicoccin, which stimulates H+-ATPase activity, also stimulated BNIs release in the absence of NH4+; vanadate, which suppresses H+-ATPase activity, also suppressed the release of BNIs. NH4+ levels (in rhizosphere) positively influenced BNIs release and root H+-ATPase activity in the concentration range of 0-1.0 mM, indicating a close relationship between BNI release and root H+-ATPase activity with a possible involvement of carrier-mediated transport for the release of BNIs in sorghum. Conclusion Our results suggest that NH4+ uptake, PM H+-ATPase activity, and rhizosphere acidification are functionally inter-connected with BNI release in sorghum. Such knowledge is critical to gain insights into why BNI function is more effective in light-textured, mildly acidic soils compared to other soil types.